Comparison of formation of reactive conformers for the S _N 2 displacements by CH ₃ CO\documentclass[12pt]{minimal}\usepackage{amsmath}\usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy}\usepackage{mathrsfs}\setlength{\oddsidemargin}{-69pt}\begin{document}\begin{equation*}{\mathrm{_{2}^{-}}}\end{equation*}\end{document} in water and by Asp124-CO\documentclass[12pt]{minimal}\usepackage{amsmath}\usepackage{wa

Abstract

The SN2 displacement of Cl ؊ from 1,2-dichloroethane by acetate (CH 3CO2 ؊ ) in water and by the carboxylate of the active site aspartate in the haloalkane dehalogenase of Xanthobacter autothropicus have been compared by using molecular dynamics simulations. In aqueous solution, six families of contact-pair structures (I-VI) were identified, and their relative concentrations and dissociation rate constants were determined. The near attack conformers (NACs) required for the S N2 displacement reaction are members of the IV (CH 3COO ؊ ⅐ ⅐ ⅐CH2(Cl)CH2Cl) family and are formed in the sequence II3III3IV3 NAC. The NAC subclass is defined by the OCOO ؊ ⅐ ⅐ ⅐COCl contact distance of <3.41 Å and the OCOO ؊ ⅐ ⅐ ⅐COCl angle of 157-180°. The mole percentage of NACs is 0.16%, based on the 1 M standard state. This result may be compared with 13.4 mole percentage of NACs in the Michaelis complex in the enzyme. It follows that NAC formation in the enzyme is favored by 2.6 kcal͞mol. Because reaction coordinates from S to TS, both in water and in the enzyme, pass via NAC (i.e., S 3 NAC 3 TS), the reduction in the S 3 NAC barrier by 2.6 kcal͞ mol accounts for Ϸ25% of the reduction of total barrier in the S 3 TS (10.7 kcal͞mol). The remaining 75% of the advantage of the enzymatic reaction revolves around the efficiency of NAC 3 TS step. This process, based on previous studies, is discussed briefly. Xanthobacter autothropicus haloalkane dehalogenase (DhlA) catalyzes the S N 2 displacement of the halogen substituent from haloalkanes by Asp-CO 2 Ϫ . This reaction can be compared with the reaction in water with CH 3 CO 2 Ϫ (AcO Ϫ ) as a nucleophile (Scheme 1). The activation barrier (⌬G ) for the displacement of Cl Ϫ from dichloroethane (DCE) is 15.3 kcal͞mol for the enzymatic reaction (1) and Ϸ26 kcal͞mol for the nonenzymatic counterpart in water (2). The first goal of this study is to devise means of determining the time-dependent mechanism of forming contact pairs from two separate reactants in water that satisfy the structural restraints of a near attack conformation (NAC). The second objective is to evaluate the contribution of NAC formation to the kinetic advantage of the DhlA reaction over the model reaction in water. These procedures can be generally applied to a study of various enzymatic catalysis over a water reaction. Methods Molecular Dynamics (MD) Simulation of 1,2-DCE and Acetate in Water.To compare ground state structures of reactants in water with those in the enzyme active site, an MD simulation was performed on the system consisting of 1,2-DCE and an acetate ion (AcO Ϫ ) in a box of TIP3P water (3). Standard CHARMM (version 27) force field was used for AcO Ϫ although OPLS force field parameters were adopted for DCE. The torsional parameters for DCE were adjusted so that only one of the two gauche conformations was sampled. It is known that for the S N 2 displacement of Cl Ϫ , DCE should be in a gauche conformation (4-6). After the reactants were placed in a water box of size 20 ϫ 20 ϫ 20 Å 3 , any TIP3P water molecu...

Stepwise dissection and visualization of the catalytic mechanism of haloalkane dehalogenase LinB using molecular dynamics simulations and computer graphics

“…In common with reports for another dehalogenase, we consider that for NAC stabilization the following three conditions need to be simultaneously fulfilled: (i) a distance between the nucleophilic oxygen (OD1) of Asp108 and the attacked carbon of DBE (C2) 3.4 Å , (ii) an angle between OD1 of Asp108, C2 and the leaving halogen between 1578 and 1808 [36], and (iii) adequate stabilization of the leaving halogen by hydrogen bonding interactions with the side chain nitrogens of Asn38 and Trp109 which assist in formation of the halide by further polarizing the C 2 -Br 1 covalent bond.…”

Molecular basis for the high-affinity binding and stabilization of firefly luciferase by PTC124

“…However, in the active site of the enzyme, the correct "near-attack conformers" (NACs) of the reactants are brought within van der Waals distance allowing bond formation, presumably via S N 2 displacement (Fig. 3A) (23).…”

Mechanisms and Free Energies of Enzymatic Reactions

“…47,116,169,217,220,[223][224][225][226][227][228][229] Bruice and co-workers have focused on identifying enzyme configurations that are most suitable for the nucleophilic attack. 116,227 However, Shurki et al suggested that these near attack conformations do not make important contributions to catalysis. 220 An enzyme that employs both transition state stabilization and reactant state destabilization, induced by substrate binding, is chorismate mutase, which catalyzes what is formally a Claisen rearrangement of chorismate to prephenate in the biosynthesis of aromatic residues.…”